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The epidermis is a stratified squamous epithelium in which keratinocytes progressively undergo terminal differentiation towards the skin surface leading to programmed cell death. In this respect we studied the role of caspases. Here, we show that caspase-14 synthesis in the skin is restricted to differentiating keratinocytes and that caspase-14 processing is associated with terminal epidermal differentiation. The proapoptotic executioner caspases-3, -6, and -7 are not activated during epidermal differentiation. Caspase-14 does not participate in apoptotic pathways elicited by treatment of differentiated keratinocytes with various death-inducing stimuli, in contrast to caspase-3. In addition, we show that non-cornifying oral keratinocyte epithelium does not express caspase-14 and that the parakeratotic regions of psoriatic skin lesions contain very low levels of caspase-14 as compared to normal stratum corneum. These observations strongly suggest that caspase-14 is involved in the keratinocyte terminal differentiation program leading to normal skin cornification, while the executioner caspases are not implicated. [ABSTRACT FROM AUTHOR]

The article discusses the histopathological changes in the coronary arteries of patients with acute coronary syndromes such as unstable angina, myocardial infarction and sudden ischemic death. Important factors in the development of acute coronary syndromes are the destabilization of atherosclerotic plaque and the superimposed formation of the platelet-rich thrombus.

Infantile haemangioma (IH) may present as a precursor area of pallor prior to the initial proliferative phase, which implies that the early lesion may be hypoxic. To examine the effect of hypoxia on the expression and activity of two key molecular markers of IH, glucose transporter-1 (GLUT1) and indoleamine 2,3-dioxygenase (IDO). IH endothelial cells express both haematopoietic and endothelial cell markers. CD14+ monocyte-derived endothelial-like cells have been employed in the study of IH and is the cell type used in this study. GLUT1 transcript, protein and activity levels were strongly induced by hypoxia and remained elevated following 2 days of normoxic recovery. IDO transcript levels were not affected by hypoxia, although IDO protein level was reduced fivefold and IDO activity > 100-fold following 2 days of hypoxia. The protein and activity levels returned to normal following 2 days of normoxic recovery. The findings link the tissue hypoxia that precedes lesion development and the expression and/or activity of two key IH proteins. The early hypoxic insult may contribute to the elevated GLUT1 levels in IH lesions, while the very low IDO activity during the hypoxic phase may promote activation of immune cells in the lesion, which release cytokines that trigger IDO expression and activity and entry into the proliferative phase. Interestingly, IH lesion development shares some common features with ischaemia-reperfusion injury. [ABSTRACT FROM AUTHOR]

The destruction of cartilage is an important characteristic of rheumatoid arthritis (RA). Immune complexes (IC) are usually found in high amounts in RA synovial fluids (SF) and in the superficial layers of RA cartilage. The objective of this study was to investigate if IC have a direct influence on proliferation, survival and production of nitric oxide (NO) of cytokine-activated chondrocytes. Primary bovine chondrocytes were incubated with cytokines (huIL-1α, bovIFN-γ, huTNF-α) and IC containing precipitates of peripheral blood (PB) and/or synovial fluid (SF) of 14 RA patients, 5 osteoarthritis (OA) patients and 10 healthy age and sex-matched controls. After 48 h, chondrocyte viability, proliferation, apoptosis, NO production and oxygen radical levels were measured. Staining with May-Grünwald-Giemsa after incubation with IC of RA PB and SF, showed apoptotic chondrocytes with condensation of the nuclei. The proliferation rates of cytokine-activated chondrocytes, incubated with sera and SF IC of RA patients were significantly decreased compared to chondrocytes, incubated with sera and SF IC of OA patients and compared to sera of controls. Quantitative evaluation of apoptotic cells by annexin-V/propidium iodide and TUNEL assays revealed a significant increase after incubation with sera and SF IC of RA patients, compared to control sera and OAs sera and SF. In all TUNEL positive samples, active-caspase-3-positive cells were found. There was a significant increase of chondrocyte NO production, after incubation with SF IC of RA patients, compared to OA SF. These results support the hypothesis that IC, present in serum and SF of RA patients, have a profound influence on chondrocyte growth, NO production and apoptosis, contributing to cartilage destruction in RA. [ABSTRACT FROM AUTHOR]

Our aim was to investigate the relationship between the various histopathological features and the CT and MRI findings in routinely submitted histopathological specimens for the diagnosis of tuberculous lymphadenopathy. Twelve formalin-fixed, paraffin-embedded tissue blocks from ten patients who were clinically suspected of having tuberculous lymphadenopathy were evaluated. We assessed the presence of histopathological features including granuloma formation, caseous necrosis, and presence of Langhans-type giant cells, calcifications, fibrosis or normal lymphoid tissue. We performed polymerase chain reaction (PCR)-based assay for mycobacterial DNA and Ziehl-Neelsen staining for acid-fast bacilli (AFB). Findings were compared with those of CT and MRI, including signal intensities on unenhanced MR images, lymph node homogeneity, attenuation values on contrast-enhanced CT and enhancement patterns on MRI. Based on CT and MRI findings, four lymph node types could be defined: (1) homogeneous nodes, visible on both pre- and post-contrast images and corresponding histopathologically to granulation tissue without or with minimal caseation necrosis (n = 2); (2) heterogeneous nodes, showing heterogeneous enhancement patterns with central non-enhancing areas and corresponding to minor or moderate intranodal caseation/liquefaction necrosis (n = 3); (3) nodes showing peripheral rim enhancement and corresponding to moderate or extensive intranodal caseation/liquefaction necrosis (n = 5); (4) heterogeneous nodes showing intranodal hyperdensities on CT and hypointense areas on T1- and T2-weighted images and corresponding to fibrosis and calcifications (n = 2). On CT and MRI, the findings reflect different stages of the tuberculous process. Imaging findings depend on the presence and the degree of granuloma formation, caseation/liquefaction necrosis, fibrosis and calcifications. [ABSTRACT FROM AUTHOR]

Paradoxical clinical deterioration of miliary tuberculosis, characterized by pulmonary and abdominal manifestations, is reported in a patient with the acquired immunodeficiency syndrome, after initiation of treatment with highly active antiretroviral therapy. Paradoxical reaction was attributed to partial restoration of cell-mediated immunity related to highly effective antiretroviral therapy. Because tuberculosis has a high prevalence in HIV patients and tuberculosis is often characterized by miliary spreading of disease in these patients, it is important to recognize this phenomenon. [ABSTRACT FROM AUTHOR]

Reactive oxygen species (ROS)-induced DNA damage has recently been identified in both human and experimental atherosclerosis. This study was undertaken to investigate whether RNA damage occurs in human atherosclerotic plaques and whether this could be related to oxidative stress. The integrity of total RNA isolated from carotid endarterectomy specimens ( n = 20) and nonatherosclerotic mammary arteries ( n = 20) was analyzed using an Agilent 2100 Bioanalyser (Agilent Technologies, Palo Alto, CA). Oxidative modifications of RNA were detected by immunohistochemistry. Eleven out of 20 atherosclerotic plaques showed a significant reduction of the 18S/28S rRNA peaks and a shift in the RNA electropherogram to shorter fragment sizes. In contrast, all mammary arteries showed good-quality RNA with clear 18S and 28S rRNA peaks. Strong nuclear and cytoplasmic immunoreactivity for oxidative damage marker 7,8-dihydro-8-oxo-2′-guanosine (8-oxoG) could be detected in the entire plaque in smooth muscle cells (SMCs), macrophages and endothelial cells, but not in SMCs of adjacent normal media or in mammary arteries. Cytoplasmic 8-oxoG staining in the plaque clearly diminished when tissue sections were pretreated with RNase A, suggesting oxidative base damage of RNA. In vitro treatment of total RNA with ROS-releasing compounds induced RNA degradation. Both loss of RNA integrity and 8-oxoG oxidative modifications were found in human atherosclerotic plaques. Because RNA damage may affect in vitro transcript quantification, RT-PCR results must be interpreted cautiously if independent experimental validation (e.g. evaluation of RNA integrity) is lacking. Eur J Clin Invest 2004; 34 (5): 323–327 [ABSTRACT FROM AUTHOR]

For the first time estrogen DNA-adducts were identified in DNA human breast tumor tissue using nano-LC coupled to nano-Electrospray Tandem Mass Spectrometry. Normal breast tissue was analyzed analogously. The data obtained in the five breast tumor and five adjacent normal tissue samples were compared qualitatively, but no straightforward difference was observed. Prior to LC-MS analysis the DNA was enzymatically hydrolyzed to a nucleoside pool. The DNA-hydrolysates were directly injected onto a column switching system developed for on-line sample clean-up and subsequent analysis of the DNA-adducts. In four patients using Premarin, DNA-adducts of 4-hydroxy-equilenin (4OHEN) were detected. All except three samples contained DNA-adducts from 4-hydroxy-estradiol or 4-hydroxy-estrone. Also DNA isolated from eight alcohol fixed and paraffin embedded breast tumor tissue showed the presence of different estrogen DNA-adducts. Worthwhile mentioning is the presence of adducts responding to m/z 570 > m/z 454 transition. This is a well-known SRM-transition indicative for the presence of the 2′-deoxyguanosine (dGuo) adduct of Benzo[a]pyrene. [Copyright &y& Elsevier]

A 71-year-old patient was referred for suspected hyperthyroidism because of a 15-kg weight loss, suppressed thyroid stimulating hormone (TSH), and a 4-cm nodule in the left thyroid lobe. Both free T4 and T3 were normal. Antithyroglobulin, anti-TSH receptor and antimicrosomal antibodies were absent. Thyroid scintigraphy showed a cold nodule in the left thyroid lobe. CAT scan of the neck revealed a 4-cm inhomogeneous nodule at the left side. An elevated sedimentation rate suggested bacterial thyroiditis, localized Quervain thyroiditis, malignancy, and the fibrosing variant of Hashimoto’s thyroiditis or Riedel’s thyroiditis. A fine needle biopsy of the thyroid nodule showed no malignant cells but was inconclusive. A true cut biopsy demonstrated atypical inflammation and also failed to reveal the diagnosis. Therefore, the patient was admitted to the hospital for further work-up and was unexpectedly found to have nodular lesions in the lung on a chest X-ray. Additional blood analysis revealed a positive cytoplasmic ANCA-titer. After inconclusive peripheral lung biopsies, a left hemithyroidectomy and a very large video-assisted thoracoscopic lung biopsy were performed, both revealing extensive zones of necrosis surrounded by granulomatous foci pointing to the diagnosis of Wegener’s granulomatosis (WG) disease. To our knowledge, this is the first report of a well-documented WG of the thyroid gland. Although extremely rare, WG should be included in differential diagnosis of inflammatory lesions of the thyroid gland. [ABSTRACT FROM AUTHOR]

Several lines of evidence suggest that macrophages play a key role in atherosclerotic plaque destabilization and rupture. Selective removal of macrophages from plaques via pharmacological therapy could therefore represent a promising approach to stabilize ‘vulnerable’, rupture-prone lesions. Yet, how macrophages can be eliminated from plaques without influencing other cell types, including smooth muscle cells (SMCs), is unknown. In the present study, we report that benzyloxycarbonyl-Val-Ala-DL-Asp(O-methyl)-fluoromethylketone (z-VAD-fmk), a caspase inhibitor with broad specificity, induces nonapoptotic cell death of macrophages, but not of SMCs. Cell death was characterized by bulk degradation of long-lived proteins, processing of microtubule associated protein light chain 3, and cytoplasmic vacuolization, which are all markers of autophagy. However, also necrosis occurred and the number of necrotic cells rapidly increased during z-VAD-fmk treatment. Western blots revealed that receptor interacting protein 1 (RIP1) is less abundantly expressed in SMCs as compared with macrophages, which may explain the different sensitivity to z-VAD-fmk. Selective degradation of RIP1 in macrophages by geldanamycin protected cells against z-VAD-fmk induced death, whereas overexpression of RIP1 in SMCs stimulated z-VAD-fmk mediated cell death. Importantly, upon z-VAD-fmk treatment macrophages overexpressed and secreted several chemokines and cytokines, including tumor necrosis factor-α (TNFα). The combination of z-VAD-fmk and TNFα but not TNFα alone, induced SMCs necrosis via a mechanism that required RIP1 expression. These results suggest that z-VAD-fmk, despite its selective cell death inducing capacity, would be detrimental for the stability of atherosclerotic plaques due to enlargement of the necrotic core, stimulation of inflammatory responses and indirect induction of SMC death. [ABSTRACT FROM AUTHOR]